The most common form of dementia among the elderly is Alzheimer's disease. Alzheimer's disease is a progressive, neurodegenerative disease characterized by memory loss, language deterioration, impaired visuospatial skills, poor judgment, indifferent attitude, but preserved motor function. At the neuroanatomical level, Alzheimer's disease is characterized by loss of synapses and neuronal cell death accompanied by the formation of extracellular senile plaques and intracellular neurofibrillary tangles.
The causes of Alzheimer's disease have been widely studied, but remain poorly understood. Nevertheless, the initial molecular events in Alzheimer's disease resulting in neuronal cell death are widely considered to involve senile plaques. The senile plaques in the diseased brain are irregular, approximately spherical, and are found commonly in the cerebral cortex and hippocampus of brains of Alzheimer's disease patients. A major component of the senile plaques is a form of amyloid peptide. The amyloid peptide that accumulates in the senile plaques notably contains β-pleated sheets, whereas the normal amyloid peptide contains α-helices and random coils.
The particular beta amyloid (Aβ) peptide that accumulates in senile plaques is a 39-43 amino acid degradation product of the naturally-occurring transmembrane protein amyloid precursor protein. Soluble a-helical or random coil conformations of Aβ peptides have little or no neurotoxicity. However, in vitro studies have established that the fibrillar β-pleated sheet conformation of Aβ is neurotoxic (See, e.g., Yankner et al., Science 1990; 250: 279-282, the contents of which are incorporated by reference). One of the significant questions in Alzheimer's disease research is, Why does Aβ peptide begin to form β-pleated sheets to such a large degree in Alzheimer's disease?
Membrane phospholipid changes in addition to abnormal Aβ formation have been reported in Alzheimer's disease. Research has demonstrated major alterations in membrane phospholipid metabolism in the brains of Alzheimer's disease patients, including changes in phospholipid composition (Pettegrew et al., Neurochem. Res. 2001; 26: 771-782, the contents of which are hereby incorporated by reference); changes in phospholipid metabolic enzymes (See, e.g., Kanfer et al., Neurochem. Res. 1993; 18: 331-334, the contents of which are incorporated herein by reference); and changes in the precursors and breakdown products of membrane phospholipids (See, e.g., Pettegrew et al., Brain Res. Bull. 2000, 53(4): 455-469, the contents of which are incorporated herein by reference). Changes in membrane phospholipid and high-energy phosphate metabolism have been demonstrated by in vivo 31P magnetic resonance imaging in a pre-symptomatic individual 33 months prior to the diagnosis of possible incipient dementia and 46 months prior to the diagnosis of Alzheimer's disease dementia. Together, these data suggest a causal role of membrane phospholipid metabolic processes in the development of Alzheimer's disease.
Glycerophosphocholine (C8H20PO6N, hereinafter abbreviated as GPC) is a normal membrane phospholipid breakdown product of phosphatidylcholine that is produced by the combined action of phospholipase A and lysophospholipase activity in all tissues, including brain. The normal levels of GPC in adult human brain are 1-2 mM and the levels are developmentally and aging regulated. The levels of GPC in brain naturally increase with age, but increase to a much greater degree in patients with Alzheimer's disease. The parallels between brain GPC levels and Alzheimer's disease raises the intriguing possibility that GPC plays a causal role in Alzheimer's disease.
In vitro studies were conducted to examine if GPC could affect the formation of Aβ peptide β-sheet deposits. The results of those studies showed that GPC enhanced Aβ(1-40) aggregation by over 400% (see, e.g., Klunk et al., J. Neurochem. 1997; 69: 266-272, the contents of which are incorporated herein by reference). Further studies demonstrated that GPC reduces the α-helical content of Aβ(1-40) by 15%, thus directly demonstrating that GPC is able to alter the conformation of Aβ peptides (See, e.g., Pettegrew et al., 2003 Abstract Viewer, Soc. Neurosci., Program No. 944.2, the contents of which are incorporated herein by reference). These results further support the idea that GPC may play a causal role in the development of Alzheimer's disease.
Computer-based molecular modeling studies have suggested a mechanism of action by which GPC stabilizes a β-turn in Aβ peptide. The molecular interaction of GPC with Aβ(1-28), which is known to determine the kinetics of Aβ aggregation, was studied by computer-based molecular mechanics and dynamics modeling (See, e.g., McClure et al., Soc. Neurosci. Abstr. 2001; 27, abstract 322.9, the contents of which are incorporated herein by reference). GPC was found to bind specifically to a site in the Aβ(1-28) peptide that forms a pocket comprised of three amino acids—namely Lys28, -Asp23, and -Lys16. Moreover, GPC binding to this peptide pocket promotes a transition β-turn conformation of the Aβ peptide. Introducing a β-turn in the Aβ peptide is critical and essential in order for Aβ peptide to form a β-sheet conformation which is the Aβ conformation which aggregates into senile plaques (FIG. 1). These data pointed to GPC as possibly playing a causal role in β-sheet aggregation of Aβ peptide.
To address the question of the specificity of GPC binding to Aβ peptides, protein structures deposited in the Protein Data Bank (PDB) were examined. In this molecular modeling approach, proteins were inspected for potential binding sites of GPC that resembled the binding site on Aβ peptides. Some 11,996 structures were compared and the top 117 matches were examined further to determine the matches that contained the correct sequence. Protein homology analysis further limited the results to five likely candidates. However, visual inspection of these protein structures did not reveal binding pockets similar to the proposed site for GPC. Based on this search and additional searches of the PDB database, it appears that the Lys28-Asp23-Lys16 motif that produces the proposed GPC binding site is unique to Aβ peptide structures. Furthermore, the spatial orientation of the Lys28-Asp23-Lys16 residues found in Aβ also appears to be important. These observations suggest that the binding of GPC to Aβ is highly specific.
Since both GPC and Aβ interact with or are generated from the cellular lipid membrane, the manner in which these two molecular species might normally interact was investigated. Fluorescence spectroscopy studies were conducted on normal human erythrocytes and rat brain membranes and results from them confirmed that GPC and Aβ(1-40) interact within the natural environment of membrane phospholipids (See, e.g., Mandal et al., submitted to Neurochem. Res. 2004, the contents of which are incorporated herein by reference).
Previous research efforts have demonstrated that GPC can induce β turns in the Aβ peptide, which may then lead to the formation of extracellular β sheets. The β sheets may, in turn, promote the formation of aggregates of Aβ peptide in senile plaques in Alzheimer's disease, leading to loss of synapses and neuronal cell death. In Alzheimer's disease, brain concentrations of GPC have been shown to increase with age and prior to the expression of Alzheimer's disease symptoms.
However, there has been no suggestion or teaching in the prior art of how to employ these scientific observations in the treatment or diagnosis of Parkinson's disease.
There is also a long-standing need within the medical community for a diagnostic tool for assessing the pre-symptomatic onset of Parkinson's disease. Thus, it is desirable to provide compositions and methods that are useful for the pre-symptomatic measurement of in the development of Parkinson's disease.